Organ transplants of liver, kidney, heart, and lung are now regularly performed now as treatment for end-stage organ disease. Allograft (same species donor and recipient) as well as xenograft (different species donor and recipient) transplants have been performed. Two primary problems for all organ transplants, however, have been acute rejection of the donor organ and the high risk of infection. Treatment for acute rejection, intensification of immunosuppression, causes deterioration of immune function, resulting in increased susceptibility to serious infection.
Organ transplants evoke a variety of immune responses in the recipient. In acute rejection, the graft is initially invaded by recipient mononuclear cells (macrophage, lymphocyte and monocyte cells). If these cells perceive antigenic differences in the graft, they will process and present the antigen to a T-lymphocyte and activate it in an antigen-specific manner. The T-cell then stimulates the central lymphoid system to elicit an immune response. The response is usually a combination of cellular (T-cell mediated) and humoral (B-cell mediated) responses. The former reaction appears to be the primary cause of the initial acute transplant rejection occurring one to three weeks post-transplant. The outcome of this acute rejection depends in part on whether immunosuppressive treatment is effective.
Acute rejection is reported to occur in 50 to 70% of hepatic grafts, depending on the criteria for diagnosis. Although quite common, few transplanted livers fail because of uncontrollable acute rejection. Relative ease of acute rejection control of hepatic grafts is not seen with other solid organ grafts, such as kidney, pancreas, and cardiac grafts.
In liver transplant patients, acute rejection is characterized by two consecutive days of rising bilirubin or liver enzymes (such as SGOT, SGPT, and alkaline phosphatase), which would indicate graft dysfunction, and simultaneous histologic findings of rejection on biopsy. The earliest histologic changes characteristic of acute rejection are accumulation of mononuclear cells in the portal tracts. The infiltrate consists of lymphocytes, and to a lesser extent neutrophils and eosinophils. Infiltrates that spill over into the parenchyma constitute a more specific sign of established acute rejection. The presence of eosinophils and the polymorphonuclear cells (PMNs) is often obscured by a prominent lymphocytic infiltrate. Eosinophilia and endothelialitis of the central vein and portal vein are also seen. Histologic evidence of biliary damage is reported to occur in 10 to 75% of patients with acute rejection. See Foster et al., Transplantation, 47:72-74 (1989) and Williams et al., Seminars in Liver Disease, 12:60-72 (1992).
In heart transplant patients, acute rejection is characterized by clinical signs of fever, arrhythmia, congestive heart failure, and increased cardiac volumes on echocardiogram. The diagnosis is established by transvenous endomyocardial biopsy using grading criteria published by Billingham, J. Heart Transplant, 9:272-276 (1990).
In lung transplant patients, acute rejection is characterized by clinical signs of fever, leukocytosis, bronchorrhea, and increasing alveolar to arteriolar oxygen gradient, all in the absence of pulmonary infection. Radiographic findings on chest X-ray may be normal or may show bi-perihilar infiltrates. Spirometry typically shows decreased forced expiratory volume over one second. The final diagnosis is established on clinical grounds, by response to bolus steroids, and on the basis of transbronchial biopsy. These criteria have been discussed in Paradis et al., J. Heart and Lung Transplant, 11:S232-6 (1992).
In kidney transplant patients, acute rejection is characterized by deteriorating renal function as shown by increasing BUN and creatinine, graft enlargement, fever, oliguria, hypertension, and reduced renal clearances. Renal scans will initially show a reduction in excretion with cortical retention, followed in several days by reductions in cortical uptake as well. If the rejection episode occurs during a period of acute tubular necrosis, its diagnosis may be delayed, being made either by serial scan assessment or by a transplant biopsy during a febrile episode. Lymphocymria is often found and may be helpful, along with a negative urine culture in ruling out graft pyelonephritis. Renal biopsies performed at this time typically reveal interstitial nephritis, mononuclear cell infiltrate, acute arteritis, and glomerular injury. Patients with multiple or severe early rejections have worse graft functional outcomes (at one, two, and five years) than patients without.
Granulocyte colony stimulating factor (G-CSF) is one of the hematopoietic growth factors, also called colony stimulating factors, that stimulate committed progenitor cells to proliferate and to form colonies of differentiating blood cells, G-CSF preferentially stimulates the growth and development of neutrophils, and is useful for treating in neutropenic states. Welte et al., PNAS-USA 82: 1526-1530 (1985); Souza et al., Science 232: 61-65 (1986) and Gabrilove, J. Seminars in Hematology 26: (2) 1-14 (1989). G-CSF increases the number of circulating granulocytes and has been reported to ameliorate infection in sepsis models. G-CSF administration also inhibits the release of tumor necrosis factor (TNF), a cytokine important to tissue injury during sepsis and rejection. See, e.g., Wendel et al., J. Immunol., 149:918-924 (1992).
In humans, endogenous G-CSF is detectable in blood plasma. Jones et al., Bailliere's Clinical Hematology 2 (1): 83-111 (1989). G-CSF is produced by fibroblasts, macrophages, T cells trophoblasts, endothelial cells and epithelial cells and is the expression product of a single copy gene comprised of four exons and five introns located on chromosome seventeen. Transcription of this locus produces a mRNA species which is differentially processed, resulting in two forms of G-CSF mRNA, one version coding for a protein of 177 amino acids, the other coding for a protein of 174 amino acids. Nagata et al., EMBO J 5: 575-581 (1986). The form comprised of 174 amino acids has been found to have the greatest specific in vivo biological activity. G-CSF is species cross-reactive, such that when human G-CSF is administered to another mammal such as a mouse, canine or monkey, sustained neutrophil leukocytosis is elicited. Moore et al. PNAS-USA 84: 7134-7138 (1987).
Human G-CSF can be obtained and purified from a number of sources. Natural human G-CSF (nhG-CSF) can be isolated from the supernatants of cultured human rumor cell lines. The development of recombinant DNA technology has enabled the production of commercial scale quantities of G-CSF in glycosylated form as a product of eukaryotic host cell expression, and of G-CSF in non-glycosylated form as a product of prokaryotic host cell expression. See, e.g., U.S. Pat. No. 4,810,643 (Souza) incorporated herein by reference.
G-CSF has been found to be useful in the treatment of indications where an increase in neutrophils will provide benefits. For example, for cancer patients, G-CSF is beneficial as a means of selectively stimulating neutrophil production to compensate for hematopoietic deficits resulting from chemotherapy or radiation therapy. Other indications include treatment of various infectious diseases and related conditions, such as sepsis, which is typically caused by a metabolite of bacteria. G-CSF is also useful alone, or in combination with other compounds, such as other cytokines, for growth or expansion of cells in culture for example, for bone marrow transplants. G-CSF has been administered to transplant patients as an adjunct to treatment of infection or for treatment of neutropenia. See Diflo et al., Hepatology, 16:PA278 (1992), Wright et al., Hepatology, 14:PA48 (1991), Lachaux ell al., J. Pediatrics, 123:1005-1008 (1993), and Colquhoun et al., Transplantation, 56:755-758 (1993).